Real-Time Oligonucleotide Sequence Confirmation - METTLER TOLEDO
On Demand Webinar

Real-Time Oligonucleotide Sequence Confirmation

On Demand Webinar

Process Control, Automation, PLS-Discriminant Analysis and Validation with Inline ATR-FTIR

Real-Time Oligonucleotide Sequence Confirmation
Real-Time Oligonucleotide Sequence Confirmation

Nucleotide phosphoramidites (henceforth referred to as “amidites”) are used as building blocks during Antisense Oligonucleotide (ASO) synthesis. Before the start of ASO synthesis, amidite solution lines are primed. Synthesis is then conducted by flowing a volume of a specific amidite solution through a solid-phase support, and the process is repeated using the same or different amidite solutions to construct the antisense oligonucleotide of interest.

To ensure the addition of the correct amidite solution, engineering controls such as the use of dedicated, labeled solution tanks and process inlets are in place. Program-specific recipes are employed to execute the intended sequence. However, these controls do not prevent inadvertent errors associated with preparation of amidites in the incorrect tank or execution of an incorrect process recipe. Therefore, real-time identification of incoming amidite solutions is desired during synthesizer priming and synthesis to confirm amidite solution preparation/connection and oligonucleotide sequence identification (ID), respectively. Biogen identified the opportunity of real-time amidite solution identification using inline Attenuated Total Reflectance (ATR) Fourier Transform Infrared (FTIR) spectroscopy. ATR-FTIR offered the fundamental capability of identifying components in simple matrices in real-time and in-situ when processing ATR-FTIR outputs with a Partial Least Squares Discriminate Analysis (PLS-DA) classification model.

This presentation shares Biogen’s experience developing the PLS-DA classification method, designing the commercial PAT application and implementing the automated system.

Dan Hill - Biogen
English

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